Lee Sang-Hoon, Ahsan Nagib, Lee Ki-Won, Kim Do-Hyun, Lee Dong-Gi, Kwak Sang-Soo, Kwon Suk-Yoon, Kim Tae-Hwan, Lee Byung-Hyun
Major of Dairy Science, Division of Applied Life Science (BK21), PMBBRC, College of Agriculture and Life Sciences, Gyeongsang National University, Jinju 660-701, Korea.
J Plant Physiol. 2007 Dec;164(12):1626-38. doi: 10.1016/j.jplph.2007.01.003. Epub 2007 Mar 13.
To mitigate the oxidative damage inflicted by biotic or abiotic stresses, plants have evolved complex anti-oxidative defense mechanisms that involve induction of antioxidant and anti-oxidative enzymes, such as superoxide dismutase (SOD) and ascorbate peroxidase (APX). To determine whether overexpression of the genes encoding copper-zinc SOD (CuZnSOD) and APX in plants is capable of decreasing reactive oxygen species (ROS) produced in response to abiotic stresses, we generated transgenic tall fescue plants expressing the CuZnSOD and APX genes in chloroplasts under the control of the oxidative stress-inducible promoter, sweet potato peroxidase anionic 2 (SWPA2). Transgenic plants were generated by Agrobacterium-mediated genetic transformation, and genotypes were confirmed by DNA blot analysis. Transgenic plants were exposed to several ROS-generating abiotic stresses, such as methyl viologen (MV), H(2)O(2), and the heavy metals copper, cadmium, and arsenic, and their tolerance was evaluated. High levels of CuZnSOD and APX gene transcripts in the transgenic plants under these treatments suggested that the transgenes were functionally expressed. Compared to transgenic plants, higher amounts of ROS were generated in the leaves of control plants exposed to abiotic stresses, resulting in increased thiobarbituric acid reactive substances (TBARS), ion leakage, and chlorophyll degradation. These parameters were significantly lower in transgenic plants. Enzyme activity assays and native polyacrylamide gel electrophoresis (PAGE) showed that total SOD and APX were highly active in transgenic plants under the abiotic stresses examined. We conclude that one of the mechanisms of increased anti-oxidative defense in transgenic tall fescue plants is overexpression of the CuZnSOD and APX genes, which are utilized in scavenging ROS and thus provide improved tolerance to abiotic stresses.
为了减轻生物或非生物胁迫造成的氧化损伤，植物进化出了复杂的抗氧化防御机制，其中包括诱导抗氧化剂和抗氧化酶，如超氧化物歧化酶（SOD）和抗坏血酸过氧化物酶（APX）。为了确定植物中编码铜锌超氧化物歧化酶（CuZnSOD）和抗坏血酸过氧化物酶（APX）的基因过表达是否能够减少非生物胁迫诱导产生的活性氧（ROS），我们构建了在氧化应激诱导型启动子甘薯过氧化物酶阴离子2（SWPA2）控制下，在叶绿体中表达CuZnSOD和APX基因的转基因高羊茅植株。通过农杆菌介导的遗传转化获得转基因植株，并通过DNA印迹分析确认基因型。将转基因植株暴露于几种能产生ROS的非生物胁迫下，如甲基紫精（MV）、H₂O₂以及重金属铜、镉和砷，并评估它们的耐受性。在这些处理下，转基因植株中高水平的CuZnSOD和APX基因转录本表明转基因在功能上得到了表达。与转基因植株相比，暴露于非生物胁迫下的对照植株叶片中产生了更多的ROS，导致硫代巴比妥酸反应性物质（TBARS）增加、离子渗漏和叶绿素降解。这些参数在转基因植株中显著更低。酶活性测定和天然聚丙烯酰胺凝胶电泳（PAGE）表明，在所检测的非生物胁迫下，转基因植株中的总超氧化物歧化酶（SOD）和抗坏血酸过氧化物酶（APX）具有高活性。我们得出结论，转基因高羊茅植株抗氧化防御增强的机制之一是CuZnSOD和APX基因的过表达，它们用于清除ROS，从而提高对非生物胁迫的耐受性。
